Targeted disruption of the ubiquitously expressed nonmuscle myosin heavy chain II-B (NMHC-B) gene in mice (B-/B-) results in cardiac myocyte hypertrophy, ventricular septal defect (VSD), dextroposition of the aorta and hydrocephalus. These mice die mostly during embryogenesis. To study a less severe phenotype, we generated mice with deletion of an alternatively spliced exon and 900 bp of surrounding intron of the NMHC-B gene. Western blot analysis from various tissues of these mice (B-delta-I/B-delta-I) showed downregulation of NMHC-B. Analysis of the RNA confirmed the deletion of the exon and also showed a decrease in NMHC-B RNA. B-delta-I/B-delta-I mice were born with normal sized cardiac myocytes and did not show any evidence for structural defects. However, they developed cardiac hypertrophy with enlargement of cell size by the age of 11 months. A small percentage of these mice developed overt hydrocephalus. Pathological examination of serial brain sections showed enlargement of lateral ventricles secondary to obstruction in the area of the aqueduct of Sylvius. In order to analyze the effects of gene dosage further, we generated B-delta-I/B- mice by crossing B-delta-I/B-delta-I with B+/B- mice. B-delta-I/B- mice showed a further decrease in NMHC-B protein levels. Serial sectioning of 8 hearts from B-delta-I/B- mice revealed a VSD in 5 cases and 1 showed a transposition of the great vessels. Some of these hearts had mild to severe myocyte disarray, but measurement of the cell sizes did not show any evidence for hypertrophy. By 3-4 weeks of age, almost all B-delta-I/B- mice developed hydrocephalus and died. Pathological analysis showed similar findings that were found in B-delta-I/B-delta-I mice. Our data suggest that the extent of downregulation of NMHC-B is critical in the development of both the heart and brain defects. Together with Dr. Ferrans (NHLBI) and colleagues, we have recently identified NMHC II-B as a component of the Z-line and intercalated disks in humans and mice.